Prokaryotic expression and characterization of receptor binding domain protein of the Middle East respiratory syndrome coronavirus
10.3760/cma.j.issn.0254-5101.2016.02.004
- VernacularTitle:中东呼吸综合征冠状病毒刺突蛋白受体结合区蛋白的原核表达纯化及鉴定
- Author:
Shuai LU
;
Jiaming LAN
;
Yingzhu CHEN
;
Jianfang ZHOU
;
Kun QIN
;
Yongliang LOU
;
Wenjie TAN
- Publication Type:Journal Article
- Keywords:
Middle East respiratory syndrome;
Coronavirus;
Receptor binding domain;
Prokaryot-ic expression;
Purification;
Identification
- From:
Chinese Journal of Microbiology and Immunology
2016;36(2):98-102
- CountryChina
- Language:Chinese
-
Abstract:
Objective To express the receptor binding domain (RBD) protein of the Middle East respiratory syndrome coronavirus (MERS-CoV) and to characterize the antigenicity of the purified recombi-nant protein. Methods The codon-optimized gene encoding the RBD protein of MERS-CoV was synthesized and then cloned into the pET30a ( +) vector to construct the recombinant expression plasmid. The trans-formed E. coli BL21 (DE3) strains carrying expression plasmid were induced by IPTG under different condi-tions. The expressed products were purified by using nickel affinity chromatography and further analyzed by SDS-PAGE and Western blot assay. Indirect ELISA was performed to analyze the antigenicity and specificity of RBD proteins expressed in prokaryotic expression systems in human serological test. Results The recom-binant RBD proteins were mainly expressed as conclusion body in an optimal induction condition of 37℃ and 0. 5 mmol/ L IPTG for 4 h. The high purified recombinant RBD proteins were obtained through denaturation and renaturation with a relative molecular mass of about 29×103 . Results of the Western blot assay showed that the recombinant RBD proteins could have specific reaction with the serum samples collected form mice with MERS-CoV infection. Indirect ELISA revealed that the RBD proteins expressed in the prokaryotic ex-pression system showed better sensitivity and specificity in the detection of antibodies against MERS-CoV in human serum samples. Conclusion This study reported the prokaryotic expression and purification of RBD protein of MERS-CoV for the first time, which might pave the way for further investigation on immunological detection of MERS-CoV and development of vaccines against MERS-CoV infection.
